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The single positive T cells found in CD3-zeta/eta-/- mice overtly react with self-major histocompatibility complex molecules upon restoration of normal surface density of T cell receptor-CD3 complex.

Lin SY, Ardouin L, Gillet A, Malissen M, Malissen B - J. Exp. Med. (1997)

Bottom Line: CD3-zeta/eta-deficient mice have small thymuses containing cells that show a profound reduction in the surface levels of T cell receptors and terminate their differentiation at the CD4+CD8+ stage.Fusion of these peripheral T cells with a CD3-zeta-positive derivative of the BW5147 TCR-alpha-/beta- thymoma resulted in hybridomas that do express an heterogeneous set of T cell receptor alpha/beta dimers at their surface and at density comparable to those found in hybridomas derived from wild-type peripheral T cells.These results demonstrate that by increasing the density and/or output of the T cell receptors expressed in peripheral T cells, one can confer them with the capacity to respond to normal density of self-MHC molecules.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie, Institut National de la Sante et de la Recherche Medicale-Centre National de Recherche Scientifique de Marseille-Luminy, France.

ABSTRACT
CD3-zeta/eta-deficient mice have small thymuses containing cells that show a profound reduction in the surface levels of T cell receptors and terminate their differentiation at the CD4+CD8+ stage. Rather unexpectedly, CD3- or very low single positive T cells accumulate over time in the spleen and lymph nodes of CD3-zeta/eta-deficient mice after a process dependent on MHC expression. Fusion of these peripheral T cells with a CD3-zeta-positive derivative of the BW5147 TCR-alpha-/beta- thymoma resulted in hybridomas that do express an heterogeneous set of T cell receptor alpha/beta dimers at their surface and at density comparable to those found in hybridomas derived from wild-type peripheral T cells. We have investigated the specificities of these T cell receptors using spleen cells from congenic and mutant mouse strains, and showed that the majority of them readily recognized self-MHC class I or class II molecules. These results demonstrate that by increasing the density and/or output of the T cell receptors expressed in peripheral T cells, one can confer them with the capacity to respond to normal density of self-MHC molecules.

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Most of the CD3− peripheral single positive T cells found in  CD3-ζ/η–deficient mice display constitutively upregulated levels of  CD44. Lymph node cells from 3-mo-old CD3-ζ/η−/− and -ζ/η+/+ mice  were analyzed by three-color flow cytometry for CD4 and CD8 expression in combination with various cell surface markers. Histograms show  the expression of the markers indicated on the left on gated CD4+CD8−  cells from CD3-ζ/η−/− and -ζ/η+/+ mice. Similar patterns were observed after gating on CD4−CD8+ cells. Each histogram is compared  with a negative control histogram obtained after staining with isotypematched negative control antibodies (dotted lines).
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Figure 2: Most of the CD3− peripheral single positive T cells found in CD3-ζ/η–deficient mice display constitutively upregulated levels of CD44. Lymph node cells from 3-mo-old CD3-ζ/η−/− and -ζ/η+/+ mice were analyzed by three-color flow cytometry for CD4 and CD8 expression in combination with various cell surface markers. Histograms show the expression of the markers indicated on the left on gated CD4+CD8− cells from CD3-ζ/η−/− and -ζ/η+/+ mice. Similar patterns were observed after gating on CD4−CD8+ cells. Each histogram is compared with a negative control histogram obtained after staining with isotypematched negative control antibodies (dotted lines).

Mentions: When analyzed by flow cytometry for the expression of CD4 and CD8 molecules, thymuses from 1- to 3-mo-old CD3-ζ/η−/− mice were found to contain dramatically fewer single positive cells than age-matched wild-type thymuses (Fig. 1). Moreover, most of these cells expressed residual levels of CD25 molecules and probably corresponded to “immature” single positive cells on the way to becoming double positive cells (20). Despite the near total absence of mature single positive thymocytes, Thy1+-single positive T cells were found to accumulate over time in lymph nodes of CD3-ζ/η−/− mice (Fig. 1, right), but in smaller numbers than those from age-matched wild-type littermates (CD3-ζ/η+/+, n = 5; average 13 × 106; CD3-ζ/η−/−, n = 7, average 5.8 × 106). Similar to CD3-ζ/η−/− thymocytes, the single positive cells found in CD3-ζ/η−/− lymph nodes expressed levels of TCR/CD3 complexes barely detectable by flow cytometry (Fig. 2; 8–10, 17). When the lymph node T cell populations found in 3-mo-old wild-type and CD3-ζ/η−/− mice were compared for the expression of various cell surface markers, they both showed identical staining profiles for CD25, CD62, CD69, CD122, and Fas molecules as well as a NK1.1− phenotype (Fig. 2 and data not shown). The most striking difference was seen in the level of expression of CD44 which was highly expressed by all the single positive cells found in CD3-ζ/η−/− lymph nodes (Fig. 2). A corresponding population of Thy1+, CD3− or very low single positive T cells was also present in the spleen of 3-mo-old CD3-ζ/η–deficient mice (data not shown). It should be noted that we have not been able to extend our analysis of CD3-ζ/η−/− over a longer time because mice die around 4 mo of age after developing a chronic intestinal inflammation, characterized by diarrhea and wasting, similar to that previously documented in cytokine or TCR mutant mice (27).


The single positive T cells found in CD3-zeta/eta-/- mice overtly react with self-major histocompatibility complex molecules upon restoration of normal surface density of T cell receptor-CD3 complex.

Lin SY, Ardouin L, Gillet A, Malissen M, Malissen B - J. Exp. Med. (1997)

Most of the CD3− peripheral single positive T cells found in  CD3-ζ/η–deficient mice display constitutively upregulated levels of  CD44. Lymph node cells from 3-mo-old CD3-ζ/η−/− and -ζ/η+/+ mice  were analyzed by three-color flow cytometry for CD4 and CD8 expression in combination with various cell surface markers. Histograms show  the expression of the markers indicated on the left on gated CD4+CD8−  cells from CD3-ζ/η−/− and -ζ/η+/+ mice. Similar patterns were observed after gating on CD4−CD8+ cells. Each histogram is compared  with a negative control histogram obtained after staining with isotypematched negative control antibodies (dotted lines).
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Related In: Results  -  Collection

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Figure 2: Most of the CD3− peripheral single positive T cells found in CD3-ζ/η–deficient mice display constitutively upregulated levels of CD44. Lymph node cells from 3-mo-old CD3-ζ/η−/− and -ζ/η+/+ mice were analyzed by three-color flow cytometry for CD4 and CD8 expression in combination with various cell surface markers. Histograms show the expression of the markers indicated on the left on gated CD4+CD8− cells from CD3-ζ/η−/− and -ζ/η+/+ mice. Similar patterns were observed after gating on CD4−CD8+ cells. Each histogram is compared with a negative control histogram obtained after staining with isotypematched negative control antibodies (dotted lines).
Mentions: When analyzed by flow cytometry for the expression of CD4 and CD8 molecules, thymuses from 1- to 3-mo-old CD3-ζ/η−/− mice were found to contain dramatically fewer single positive cells than age-matched wild-type thymuses (Fig. 1). Moreover, most of these cells expressed residual levels of CD25 molecules and probably corresponded to “immature” single positive cells on the way to becoming double positive cells (20). Despite the near total absence of mature single positive thymocytes, Thy1+-single positive T cells were found to accumulate over time in lymph nodes of CD3-ζ/η−/− mice (Fig. 1, right), but in smaller numbers than those from age-matched wild-type littermates (CD3-ζ/η+/+, n = 5; average 13 × 106; CD3-ζ/η−/−, n = 7, average 5.8 × 106). Similar to CD3-ζ/η−/− thymocytes, the single positive cells found in CD3-ζ/η−/− lymph nodes expressed levels of TCR/CD3 complexes barely detectable by flow cytometry (Fig. 2; 8–10, 17). When the lymph node T cell populations found in 3-mo-old wild-type and CD3-ζ/η−/− mice were compared for the expression of various cell surface markers, they both showed identical staining profiles for CD25, CD62, CD69, CD122, and Fas molecules as well as a NK1.1− phenotype (Fig. 2 and data not shown). The most striking difference was seen in the level of expression of CD44 which was highly expressed by all the single positive cells found in CD3-ζ/η−/− lymph nodes (Fig. 2). A corresponding population of Thy1+, CD3− or very low single positive T cells was also present in the spleen of 3-mo-old CD3-ζ/η–deficient mice (data not shown). It should be noted that we have not been able to extend our analysis of CD3-ζ/η−/− over a longer time because mice die around 4 mo of age after developing a chronic intestinal inflammation, characterized by diarrhea and wasting, similar to that previously documented in cytokine or TCR mutant mice (27).

Bottom Line: CD3-zeta/eta-deficient mice have small thymuses containing cells that show a profound reduction in the surface levels of T cell receptors and terminate their differentiation at the CD4+CD8+ stage.Fusion of these peripheral T cells with a CD3-zeta-positive derivative of the BW5147 TCR-alpha-/beta- thymoma resulted in hybridomas that do express an heterogeneous set of T cell receptor alpha/beta dimers at their surface and at density comparable to those found in hybridomas derived from wild-type peripheral T cells.These results demonstrate that by increasing the density and/or output of the T cell receptors expressed in peripheral T cells, one can confer them with the capacity to respond to normal density of self-MHC molecules.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie, Institut National de la Sante et de la Recherche Medicale-Centre National de Recherche Scientifique de Marseille-Luminy, France.

ABSTRACT
CD3-zeta/eta-deficient mice have small thymuses containing cells that show a profound reduction in the surface levels of T cell receptors and terminate their differentiation at the CD4+CD8+ stage. Rather unexpectedly, CD3- or very low single positive T cells accumulate over time in the spleen and lymph nodes of CD3-zeta/eta-deficient mice after a process dependent on MHC expression. Fusion of these peripheral T cells with a CD3-zeta-positive derivative of the BW5147 TCR-alpha-/beta- thymoma resulted in hybridomas that do express an heterogeneous set of T cell receptor alpha/beta dimers at their surface and at density comparable to those found in hybridomas derived from wild-type peripheral T cells. We have investigated the specificities of these T cell receptors using spleen cells from congenic and mutant mouse strains, and showed that the majority of them readily recognized self-MHC class I or class II molecules. These results demonstrate that by increasing the density and/or output of the T cell receptors expressed in peripheral T cells, one can confer them with the capacity to respond to normal density of self-MHC molecules.

Show MeSH
Related in: MedlinePlus